Stem cell factor (SCF) is a growth factor critical for normal hematopoiesis. The receptor for SCF is c-Kit, a receptor tyrosine kinase (RTK). Activating mutations in c-Kit are associated with mastocytosis, leukemia and other cancers. Because of the importance of c-Kit in normal hematopoiesis, as well as its role in human disease, the objective of this project is to understand the mechanism of action of both normal and oncogenic forms of this receptor in hematopoietic progenitor cells and mast cells, two important target populations of SCF. One specific aim of this project is to define the biochemical mechanism of action of wild-type c-Kit. Specifically, we have focused on the role of the JAK/STAT pathway and Src family members in SCF-mediated responses. We have previously shown that SCF induces association of c-Kit and Lyn, and that Lyn activity increases at multiple points during SCF-induced cell cycle progression. This year we published work demonstrating that Lyn is required for progenitor and mast cells to respond normally to SCF. Studies defining downstream signaling pathways that couple Lyn to SCF-mediated responses are in progress. We have also shown that SCF activates the JAK/STAT pathway. JAK2 and Stat1 associate with c-Kit, and are activated by SCF. We are now investigating the biological role of this activation in SCF-mediated responses through the study of progenitor and mast cells isolated from JAK2-deficient mice. A second specific aim of this project is to delineate signal transduction pathways mediating cellular transformation by D816V c-Kit, an oncogenic mutant found in patients with mastocytosis and leukemia. This year we published work demonstrating that the D816V c-Kit mutant constitutively associates with activated PI3 kinase and that constitutive recruitment of activated PI3 kinase is essential for tumorigenicity of this mutant. Studies are in progress to determine if PI3 kinase is necessary and sufficient for transformation of hematopoietic cells. These data could have important implications in designing treatment strategies of diseases associated with this mutation, such as mastocytosis and leukemia.